Cellular Dysregulation Research Articles

Emerging contaminants at trace levels of pesticides perturbs biomolecules in different organs in mice: Role of peroxisome proliferator-activated receptor-alpha

Objectives: Information is lacking on the consequences of chronic exposure to emerging contaminants at environmentally relevant (trace concentrations) on biomolecules. Environmental exposure to these chemical mixtures happens at trace concentrations and at multiple molecular interactions. The consequences of trace concentrations of multiple pesticides (MPs) on the regulation of selected biomolecules nitric oxide (NO), thiols, superoxide dismutase (SOD), and glutathione S-transferase (GST) in the tissues from wild type (WT) and genetically deficient- peroxisome proliferator-activated receptor-alpha (PPARα) knockout (Null) mice were investigated. Material and Methods: Mice were exposed to trace concentrations of MPs: Atrazine, dieldrin, endrin, endosulfan, and anthracene (1–100 ng/L) in drinking water for 6 weeks. Organs were collected and homogenized; NO, protein and non-protein thiol levels, as well as SOD and GST activities were determined. Results: Differential and organ selective effects of the treatments were observed in the WT and PPARα knockout. Increased NO levels were observed in the organs from WT with limited increase in the kidney (Null). SOD activity was decreased in the organs from the WT and was increased in the PPARα knockout when compared to the control. Thiol level was significantly increased in the heart and spleen in the WT and in the heart of the PPARα knockout mice when compared to the control. Non-protein thiol concentration was reduced in the heart and kidney (WT) and reduced in the liver of the PPARα knockout when compared to the control. GST activity was significantly decreased in the liver and spleen (WT) and was significantly elevated in all organs in the PPARα knockout mice when compared to the WT. Conclusion: The low concentrations of MPs may have caused selective dysregulation of biomolecules in different organs of the body. These effects observed may be influenced by genetic status such as in PPARα deficiency. These results present a scenario that implicates nanoconcentrations of series of organic contaminants that can cause cellular and molecular dysregulations of biomolecules precipitating toxicity and pathology that can be a threat to human health. Further, investigation into the molecular mechanism(s) and signaling pathway(s) implicated in these dysregulations is warranted.

Chloride Intracellular Channel Protein 1 (CLIC1) Is a Critical Host Cellular Factor for Influenza A Virus Replication.

(1) Background: Influenza A Virus (IAV) uses host cellular proteins during replication in host cells. IAV infection causes elevated expression of chloride intracellular channel protein 1 (CLIC1) in lung epithelial cells, but the importance of this protein in IAV replication is unknown. (2) In this study, we determined the role of CLIC1 in IAV replication by investigating the effects of CLIC1 knockdown (KD) on IAV viral protein translation, genomic RNA transcription, and host cellular proteome dysregulation. (3) Results: CLIC1 KD in A549 human lung epithelial cells resulted in a significant decrease in progeny supernatant IAV, but virus protein expression was unaffected. However, a significantly larger number of viral RNAs accumulated in CLIC1 KD cells. Treatment with a CLIC1 inhibitor also caused a significant reduction in IAV replication, suggesting that CLIC1 is an important host factor in IAV replication. SomaScan®, which measures 1322 proteins, identified IAV-induced dysregulated proteins in wild-type cells and in CLIC1 KD cells. The expression of 116 and 149 proteins was significantly altered in wild-type and in CLIC1 KD cells, respectively. A large number of the dysregulated proteins in CLIC1 KD cells were associated with cellular transcription and predicted to be inhibited during IAV replication. (4) Conclusions: This study suggests that CLIC1 is involved in later stages of IAV replication. Further investigation should clarify mechanism(s) for the development of anti-IAV drugs targeting CLIC1 protein.

Mechanisms of Immune Dysregulation Induced by Pesticide Exposure and their Implications for Cancer: A Systematic Review

The inappropriate use of pesticides contaminates natural resources and has consequences for human health. Evidence of immune dysregulation caused by pesticides has been presented, but studies linking exposure to these substances with immune dysregulation and its relationship to cancer are scarce. This study systematically reviews the published literature that addresses the topic of immune dysregulation and cancer and its relationship to pesticide exposure. Studies developed in vitro cell culture and in vivo experimental animals and human exposure data were included. The results show that pesticide exposure has a relevant impact on immune dysfunction, negatively affecting immune system cells and cytokine production in innate and adaptive responses. The main consequences reported were suppression of the Th1 response, induction of the Th2 response, cellular balance dysregulation, and cytokine production changes. All reported events are related to the occurrence of immunosuppression. This dysregulation is paramount for cancer progression and may lead to a worse prognosis in chronically exposed populations. These findings also point to the need for further studies, primarily focused on human exposure.

Breaking barriers in trauma research: A narrative review of opportunities to leverage veterinary trauma for accelerated translation to clinical solutions for pets and people.

Trauma is a common cause of morbidity and mortality in humans and companion animals. Recent efforts in procedural development, training, quality systems, data collection, and research have positively impacted patient outcomes; however, significant unmet need still exists. Coordinated efforts by collaborative, translational, multidisciplinary teams to advance trauma care and improve outcomes have the potential to benefit both human and veterinary patient populations. Strategic use of veterinary clinical trials informed by expertise along the research spectrum (i.e., benchtop discovery, applied science and engineering, large laboratory animal models, clinical veterinary studies, and human randomized trials) can lead to increased therapeutic options for animals while accelerating and enhancing translation by providing early data to reduce the cost and the risk of failed human clinical trials. Active topics of collaboration across the translational continuum include advancements in resuscitation (including austere environments), acute traumatic coagulopathy, trauma-induced coagulopathy, traumatic brain injury, systems biology, and trauma immunology. Mechanisms to improve funding and support innovative team science approaches to current problems in trauma care can accelerate needed, sustainable, and impactful progress in the field. This review article summarizes our current understanding of veterinary and human trauma, thereby identifying knowledge gaps and opportunities for collaborative, translational research to improve multispecies outcomes. This translational trauma group of MDs, PhDs, and DVMs posit that a common understanding of injury patterns and resulting cellular dysregulation in humans and companion animals has the potential to accelerate translation of research findings into clinical solutions.

Predicting miRNA-disease association via graph attention learning and multiplex adaptive modality fusion

miRNAs are a class of small non-coding RNA molecules that play important roles in gene regulation. They are crucial for maintaining normal cellular functions, and dysregulation or dysfunction of miRNAs which are linked to the onset and advancement of multiple human diseases. Research on miRNAs has unveiled novel avenues in the realm of the diagnosis, treatment, and prevention of human diseases. However, clinical trials pose challenges and drawbacks, such as complexity and time-consuming processes, which create obstacles for many researchers. Graph Attention Network (GAT) has shown excellent performance in handling graph-structured data for tasks such as link prediction. Some studies have successfully applied GAT to miRNA-disease association prediction. However, there are several drawbacks to existing methods. Firstly, most of the previous models rely solely on concatenation operations to merge features of miRNAs and diseases, which results in the deprivation of significant modality-specific information and even the inclusion of redundant information. Secondly, as the number of layers in GAT increases, there is a possibility of excessive smoothing in the feature extraction process, which significantly affects the prediction accuracy. To address these issues and effectively complete miRNA disease prediction tasks, we propose an innovative model called Multiplex Adaptive Modality Fusion Graph Attention Network (MAMFGAT). MAMFGAT utilizes GAT as the main structure for feature aggregation and incorporates a multi-modal adaptive fusion module to extract features from three interconnected networks: the miRNA-disease association network, the miRNA similarity network, and the disease similarity network. It employs adaptive learning and cross-modality contrastive learning to fuse more effective miRNA and disease feature embeddings as well as incorporates multi-modal residual feature fusion to tackle the problem of excessive feature smoothing in GATs. Finally, we employ a Multi-Layer Perceptron (MLP) model that takes the embeddings of miRNA and disease features as input to anticipate the presence of potential miRNA-disease associations. Extensive experimental results provide evidence of the superior performance of MAMFGAT in comparison to other state-of-the-art methods. To validate the significance of various modalities and assess the efficacy of the designed modules, we performed an ablation analysis. Furthermore, MAMFGAT shows outstanding performance in three cancer case studies, indicating that it is a reliable method for studying the association between miRNA and diseases. The implementation of MAMFGAT can be accessed at the following GitHub repository: https://github.com/zixiaojin66/MAMFGAT-master.

DNA Barcoding Technology for Lineage Recording and Tracing to Resolve Cell Fate Determination.

In various biological contexts, cells receive signals and stimuli that prompt them to change their current state, leading to transitions into a future state. This change underlies the processes of development, tissue maintenance, immune response, and the pathogenesis of various diseases. Following the path of cells from their initial identity to their current state reveals how cells adapt to their surroundings and undergo transformations to attain adjusted cellular states. DNA-based molecular barcoding technology enables the documentation of a phylogenetic tree and the deterministic events of cell lineages, providing the mechanisms and timing of cell lineage commitment that can either promote homeostasis or lead to cellular dysregulation. This review comprehensively presents recently emerging molecular recording technologies that utilize CRISPR/Cas systems, base editing, recombination, and innate variable sequences in the genome. Detailing their underlying principles, applications, and constraints paves the way for the lineage tracing of every cell within complex biological systems, encompassing the hidden steps and intermediate states of organism development and disease progression.

Different Transcriptome Features of Peripheral Blood Mononuclear Cells in Non-Emphysematous Chronic Obstructive Pulmonary Disease.

Non-emphysematous chronic obstructive pulmonary disease (COPD), which is defined based on chest computed tomography findings, presented different transcriptome features of peripheral blood mononuclear cells (PBMCs) compared with emphysematous COPD. Enrichment analysis of transcriptomic data in COPD demonstrated that the "Hematopoietic cell lineage" pathway in Kyoto Encyclopedia of Genes and Genomes pathway analysis was highly upregulated, suggesting that cellular dynamic dysregulation in COPD lungs is affected by pathologically modified PBMCs. The differentially expressed genes (DEGs) upregulated in PBMCs reflected the disease state of non-emphysematous COPD. Upregulated DEGs such as XCL1, PRKCZ, TMEM102, CD200R1, and AQP1 activate T lymphocytes and eosinophils. Upregulating keratan sulfate biosynthesis and metabolic processes is associated with protection against the destruction of the distal airways. ITGA3 upregulation augments interactions with extracellular matrix proteins, and COL6A1 augments the profibrotic mast cell phenotype during alveolar collagen VI deposition. Upregulating HSPG2, PDGFRB, and PAK4 contributes to the thickening of the airway wall, and upregulating SERPINF1 expression explains the better-preserved vascular bed. Therefore, gene expression and pathway analysis in PBMCs in patients with non-emphysematous COPD represented type 2 immune responses and airway remodeling features. Therefore, these patients have asthmatic potential despite no clinical signs of asthma, in contrast to those with emphysematous COPD.

Cytokine polymorphisms in patients with autoimmune hemolytic anemia.

Autoimmune hemolytic anemia (AIHA) is due to autoantibodies with or without complement activation and involves cellular and cytokine dysregulation. Here, we investigated cytokine single-nucleotide polymorphisms (SNPs) of TNF-α, TGF-β1, IL-10, IL-6, and IFN-γ, along with their serum levels. The former were related to hematological parameters, therapy, and clinical outcome. The study included 123 consecutive patients with primary AIHA [77 warm AIHA and 46 cold agglutinin disease (CAD)], followed up for a median of 49 months. Results show that the allelic frequency of TNF-α -308 G/A polymorphisms was significantly lower in patients versus controls. Moreover, the genotypic frequency of TNF-α -308G/A and TGF-β gene codon 25 G/C genotypes was significantly lower in patients versus controls. Considering cytokine SNP genotypes associated with different gene expression levels, TNF-α high gene expression was significantly more frequent in patients, TGF-β and IL-10 high gene expression was higher in patients with more severe anemia, and TGF-β high gene expression was higher in patients with active disease. Considering treatment, TNF-α and TGF-β high gene expression was more frequent in multitreated patients and particularly in CAD. It may be speculated that this genetic predisposition to a stronger inflammatory response may result in a greater immune dysregulation and in a relapsed/refractory disease. Regarding cytokine serum levels, TNF-α and TGF-β were significantly lower, and IL-10 and IL-6 were significantly higher in patients versus controls, underlying the complex interplay between genetic background and disease features.

Feedback regulation of ubiquitination and phase separation of HECT E3 ligases

Lipid homeostasis is essential for normal cellular functions and dysregulation of lipid metabolism is highly correlated with human diseases including neurodegenerative diseases. In the ubiquitin-dependent autophagic degradation pathway, Troyer syndrome-related protein Spartin activates and recruits HECT-type E3 Itch to lipid droplets (LDs) to regulate their turnover. In this study, we find that Spartin promotes the formation of Itch condensates independent of LDs. Spartin activates Itch through its multiple PPAY-motif platform generated by self-oligomerization, which targets the WW12 domains of Itch and releases the autoinhibition of the ligase. Spartin-induced activation and subsequent autoubiquitination of Itch lead to liquid-liquid phase separation (LLPS) of the poly-, but not oligo-, ubiquitinated Itch together with Spartin and E2 both in vitro and in living cells. LLPS-mediated condensation of the reaction components further accelerates the generation of polyubiquitin chains, thus forming a positive feedback loop. Such Itch-Spartin condensates actively promote the autophagy-dependent turnover of LDs. Moreover, we show that the catalytic HECT domain of Itch is sufficient to interact and phase separate with poly-, but not oligo-ubiquitin chains. HECT domains from other HECT E3 ligases also exhibit LLPS-mediated the promotion of ligase activity. Therefore, LLPS and ubiquitination are mutually interdependent and LLPS promotes the ligase activity of the HECT family E3 ligases.

The Implications of Cannabinoid-Induced Metabolic Dysregulation for Cellular Differentiation and Growth

The endocannabinoid system (ECS) governs and coordinates several physiological processes through an integrated signaling network, which is responsible for inducing appropriate intracellular metabolic signaling cascades in response to (endo)cannabinoid stimulation. This intricate cellular system ensures the proper functioning of the immune, reproductive, and nervous systems and is involved in the regulation of appetite, memory, metabolism, and development. Cannabinoid receptors have been observed on both cellular and mitochondrial membranes in several tissues and are stimulated by various classes of cannabinoids, rendering the ECS highly versatile. In the context of growth and development, emerging evidence suggests a crucial role for the ECS in cellular growth and differentiation. Indeed, cannabinoids have the potential to disrupt key energy-sensing metabolic signaling pathways requiring mitochondrial-ER crosstalk, whose functioning is essential for successful cellular growth and differentiation. This review aims to explore the extent of cannabinoid-induced cellular dysregulation and its implications for cellular differentiation.

IgA deficiency destabilizes homeostasis toward intestinal microbes and increases systemic immune dysregulation.

The ability of most patients with selective immunoglobulin A (IgA) deficiency (SIgAD) to remain apparently healthy has been a persistent clinical conundrum. Compensatory mechanisms, including IgM, have been proposed, yet it remains unclear how secretory IgA and IgM work together in the mucosal system and, on a larger scale, whether the systemic and mucosal anti-commensal responses are redundant or have unique features. To address this gap in knowledge, we developed an integrated host-commensal approach combining microbial flow cytometry and metagenomic sequencing (mFLOW-Seq) to comprehensively define which microbes induce mucosal and systemic antibodies. We coupled this approach with high-dimensional immune profiling to study a cohort of pediatric patients with SIgAD and household control siblings. We found that mucosal and systemic antibody networks cooperate to maintain homeostasis by targeting a common subset of commensal microbes. In IgA-deficiency, we find increased translocation of specific bacterial taxa associated with elevated levels of systemic IgG targeting fecal microbiota. Associated features of immune system dysregulation in IgA-deficient mice and humans included elevated levels of inflammatory cytokines, enhanced follicular CD4 T helper cell frequency and activation, and an altered CD8 T cell activation state. Although SIgAD is clinically defined by the absence of serum IgA, the symptomatology and immune dysregulation were concentrated in the SIgAD participants who were also fecal IgA deficient. These findings reveal that mucosal IgA deficiency leads to aberrant systemic exposures and immune responses to commensal microbes, which increase the likelihood of humoral and cellular immune dysregulation and symptomatic disease in patients with IgA deficiency.

Immunological Pathways in Sarcoidosis and Autoimmune Rheumatic Disorders-Similarities and Differences in an Italian Prospective Real-Life Preliminary Study.

The pathogenesis of sarcoidosis involves T cells and B lymphocytes that produce autoantibodies. We compared the expression of different T and B cell subsets in sarcoidosis and three B-mediated rheumatic diseases that can affect the lungs in an attempt to identify similarities and differences that distinguish these diseases. The study included patients referred to Siena University Hospital's respiratory disease and rheumatology units. Patients were enrolled prospectively and consecutively. Healthy volunteers were also included. Multicolor flow cytometry was performed on phenotype T and B cell subsets. Multivariate analysis was carried out to reduce the dimensionality of the data. Fifteen patients had a diagnosis of sarcoidosis, fourteen idiopathic inflammatory myopathies (IIM), five granulomatosis with polyangiitis (GPA), ten microscopic polyangiitis (MPA), and seven were controls. Thirty-five T and B cell subsets were phenotyped, 15 of which were significantly different in sarcoidosis, B-mediated rheumatic disorders, and controls. Principal components analysis distinguished the four groups of patients with a total explained variance of 54.7%. A decision tree was constructed to determine which clustering variables would be most useful for distinguishing sarcoidosis, IIM, MPA, and GPA. The model showed regulatory T helper cells (Th-reg) > 5.70% in 91% of sarcoidosis patients as well as Th-reg ≤ 5.70 and Th17 > 43.27 in 100% of MPA. It also showed Th-reg ≤ 5.70, Th17 ≤ 43.27 and Tfh-reg ≥ 7.81 in 100% of GPA patients, and Th-reg ≤ 5.70, Th17 ≤ 43.27 and Tfh-reg ≤ 7.81 in 100% of IIM patients. The immune cell profile sheds light on similarities and differences between sarcoidosis and B-mediated rheumatic diseases. Sarcoidosis and autoimmune diseases show similar patterns of cellular immune dysregulation, suggesting a common pathogenic pathway that may provide an opportunity for further understanding autoimmunity and exploring biological therapies to treat sarcoidosis.

Single-cell atlas of the human neonatal small intestine affected by necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is a gastrointestinal complication of premature infants with high rates of morbidity and mortality. A comprehensive view of the cellular changes and aberrant interactions that underlie NEC is lacking. This study aimed at filling in this gap. We combine single-cell RNA sequencing (scRNAseq), T-cell receptor beta (TCRβ) analysis, bulk transcriptomics, and imaging to characterize cell identities, interactions, and zonal changes in NEC. We find an abundance of proinflammatory macrophages, fibroblasts, endothelial cells as well as T cells that exhibit increased TCRβ clonal expansion. Villus tip epithelial cells are reduced in NEC and the remaining epithelial cells up-regulate proinflammatory genes. We establish a detailed map of aberrant epithelial-mesenchymal-immune interactions that are associated with inflammation in NEC mucosa. Our analyses highlight the cellular dysregulations of NEC-associated intestinal tissue and identify potential targets for biomarker discovery and therapeutics.

108 Effects of Prior Hyperthyroidism Diagnosis on Burn Patient Outcomes

Abstract Introduction The metabolic response to severe burns is described by an initial hypometabolic ‘ebb’ phase and followed by a hypermetabolic ‘flow’ phase. The hypermetabolic phase is mediated by hormonal and cellular dysregulation with elevated catecholamine levels. The role of thyroid hormone, however, is not described well. This study investigates relative risks of various outcomes in those with a previous diagnosis of hyperthyroidism following severe burn compared to those without. Methods This study obtained data from the TriNetX Research Network, a national database of de-identified electronic medical data from a secure network. The population was stratified to compare patients with and without a prior diagnosis of hyperthyroidism who sustained burns. Outcomes were measured within three years of injury in order to assess long-term effects occurring alongside the hypermetabolic phase. Outcomes observed for both groups included systemic inflammation and infection, sepsis, mortality, and infections of the skin and subcutaneous tissue. Results This study identified 5,250 patients with a prior diagnosis of hyperthyroidism who sustained burn injuries and 648,336 patients without hyperthyroidism prior to matching for demographics and diagnoses. Propensity score matching then yielded 5,077 patients with and without a prior diagnosis of hyperthyroidism. Both cohorts were balanced using propensity score matching for age, sex, race, ethnicity, and TBSA burned. After assessing the outcomes for a three-year span since burn injury, patients with a prior diagnosis of hyperthyroidism had a higher risk of systemic inflammation and infection (risk ratio [RR], 1.599, p < 0.0003), sepsis ([RR], 1.572, p < 0.0001), mortality ([RR], 1.488, p < 0.0001), and developing infections of the skin and subcutaneous tissue ([RR], 1.332, p < 0.0001) compared to patients without prior diagnosis history. Conclusions Burned patients with a prior history of hyperthyroidism have increased risk of systemic inflammation and infection, sepsis, mortality, and infections of the skin and subcutaneous tissue. Thus, history of this condition or concomitant diagnosis with the injury is warranted. Applicability of Research to Practice This study encourages further insight into the relationship between the metabolic effects of burn injuries and concurrent metabolic conditions to modify patient treatment plans and customize practices.

IPSC-derived retinal pigmented epithelial cells from patients with macular telangiectasia show decreased mitochondrial function.

Patient-derived induced pluripotent stem cells (iPSCs) provide a powerful tool for identifying cellular and molecular mechanisms of disease. Macular telangiectasia type 2 (MacTel) is a rare, late-onset degenerative retinal disease with an extremely heterogeneous genetic architecture, lending itself to the use of iPSCs. Whole-exome sequencing screens and pedigree analyses have identified rare causative mutations that account for less than 5% of cases. Metabolomic surveys of patient populations and GWAS have linked MacTel to decreased circulating levels of serine and elevated levels of neurotoxic 1-deoxysphingolipids (1-dSLs). However, retina-specific, disease-contributing factors have yet to be identified. Here, we used iPSC-differentiated retinal pigmented epithelial (iRPE) cells derived from donors with or without MacTel to screen for novel cell-intrinsic pathological mechanisms. We show that MacTel iRPE cells mimicked the low serine levels observed in serum from patients with MacTel. Through RNA-Seq and gene set enrichment pathway analysis, we determined that MacTel iRPE cells are enriched in cellular stress pathways and dysregulation of central carbon metabolism. Using respirometry and mitochondrial stress testing, we functionally validated that MacTel iRPE cells had a reduction in mitochondrial function that was independent of defects in serine biosynthesis and 1-dSL accumulation. Thus, we identified phenotypes that may constitute alternative disease mechanisms beyond the known serine/sphingolipid pathway.

Post-Covid Syndrome: Pathophysiology of Systemic Disregulations

The main processes that characterize the post-COVID syndrome are analyzed as a version of the “prolonged” pathology of acute COVID-19. Considering the variety of manifestations of post-covid pathology, the main blocks of systemic, cellular and molecular dysregulations are distinguished. As the main reasons, the following are considered: the consequences of systemic and organ damage in the acute phase of COVID-19, the persistent activity of “hidden” pathogens and the altered status of the patient’s protective immune systems. Long-term pathology, as a multisystem syndrome, reflects disturbances in the main cellular and molecular regulatory systems: stochastic disorganization of immune responses, vascular endothelial dysfunction, cellular inflammation, imbalance of coagulation and antithrombosis systems, deviations in autoimmune processes, etc. These findings orient new cellular and biochemical targets for timely therapy. The varied nature of the symptoms suggests a selective use of therapeutic approaches.

Redox and metal profiles in human coronary endothelial and smooth muscle cells under hyperoxia, physiological normoxia and hypoxia: Effects of NRF2 signaling on intracellular zinc

Zinc is an important component of cellular antioxidant defenses and dysregulation of zinc homeostasis is a risk factor for coronary heart disease and ischemia/reperfusion injury. Intracellular homeostasis of metals, such as zinc, iron and calcium are interrelated with cellular responses to oxidative stress. Most cells experience significantly lower oxygen levels in vivo (2–10 kPa O2) compared to standard in vitro cell culture (18kPa O2). We report the first evidence that total intracellular zinc content decreases significantly in human coronary artery endothelial cells (HCAEC), but not in human coronary artery smooth muscle cells (HCASMC), after lowering of O2 levels from hyperoxia (18 kPa O2) to physiological normoxia (5 kPa O2) and hypoxia (1 kPa O2). This was paralleled by O2-dependent differences in redox phenotype based on measurements of glutathione, ATP and NRF2-targeted protein expression in HCAEC and HCASMC. NRF2-induced NQO1 expression was attenuated in both HCAEC and HCASMC under 5 kPa O2 compared to 18 kPa O2. Expression of the zinc efflux transporter ZnT1 increased in HCAEC under 5 kPa O2, whilst expression of the zinc-binding protein metallothionine (MT) decreased as O2 levels were lowered from 18 to 1 kPa O2. Negligible changes in ZnT1 and MT expression were observed in HCASMC. Silencing NRF2 transcription reduced total intracellular zinc under 18 kPa O2 in HCAEC with negligible changes in HCASMC, whilst NRF2 activation or overexpression increased zinc content in HCAEC, but not HCASMC, under 5 kPa O2. This study has identified cell type specific changes in the redox phenotype and metal profile in human coronary artery cells under physiological O2 levels. Our findings provide novel insights into the effect of NRF2 signaling on Zn content and may inform targeted therapies for cardiovascular diseases.

Exhaled breath condensate identifies metabolic dysregulation in patients with radiation-induced lung injury.

Radiation-induced lung injury (RILI) is a consequence of therapeutic thoracic irradiation (TR) for many cancers, and there are no FDA-approved curative strategies. Studies report that 80% of patients who undergo TR will have CT-detectable interstitial lung abnormalities, and strategies to limit the risk of RILI may make radiotherapy less effective at treating cancer. Our lab and others have reported that lung tissue from patients with idiopathic pulmonary fibrosis (IPF) exhibits metabolic defects including increased glycolysis and lactate production. In this pilot study, we hypothesized that patients with radiation-induced lung damage will exhibit distinct changes in lung metabolism that may be associated with the incidence of fibrosis. Using liquid chromatography/tandem mass spectrometry to identify metabolic compounds, we analyzed exhaled breath condensate (EBC) in subjects with CT-confirmed lung lesions after TR for lung cancer, compared with healthy subjects, smokers, and cancer patients who had not yet received TR. The lung metabolomic profile of the irradiated group was significantly different from the three nonirradiated control groups, highlighted by increased levels of lactate. Pathway enrichment analysis revealed that EBC from the case patients exhibited concurrent alterations in lipid, amino acid, and carbohydrate energy metabolism associated with the energy-producing tricarboxylic acid (TCA) cycle. Radiation-induced glycolysis and diversion of lactate to the extracellular space suggests that pyruvate, a precursor metabolite, converts to lactate rather than acetyl-CoA, which contributes to the TCA cycle. This TCA cycle deficiency may be compensated by these alternate energy sources to meet the metabolic demands of chronic wound repair. Using an "omics" approach to probe lung disease in a noninvasive manner could inform future mechanistic investigations and the development of novel therapeutic targets.NEW & NOTEWORTHY We report that exhaled breath condensate (EBC) identifies cellular metabolic dysregulation in patients with radiation-induced lung injury. In this pilot study, untargeted metabolomics revealed a striking metabolic signature in EBC from patients with radiation-induced lung fibrosis compared to patients with lung cancer, at-risk smokers, and healthy volunteers. Patients with radiation-induced fibrosis exhibit specific changes in tricarboxylic acid (TCA) cycle energy metabolism that may be required to support the increased energy demands of fibroproliferation.

Metabolic dysregulation impairs lymphocyte function during severe SARS-CoV-2 infection

Cellular metabolic dysregulation is a consequence of SARS-CoV-2 infection that is a key determinant of disease severity. However, how metabolic perturbations influence immunological function during COVID-19 remains unclear. Here, using a combination of high-dimensional flow cytometry, cutting-edge single-cell metabolomics, and re-analysis of single-cell transcriptomic data, we demonstrate a global hypoxia-linked metabolic switch from fatty acid oxidation and mitochondrial respiration towards anaerobic, glucose-dependent metabolism in CD8+Tc, NKT, and epithelial cells. Consequently, we found that a strong dysregulation in immunometabolism was tied to increased cellular exhaustion, attenuated effector function, and impaired memory differentiation. Pharmacological inhibition of mitophagy with mdivi-1 reduced excess glucose metabolism, resulting in enhanced generation of SARS-CoV-2- specific CD8+Tc, increased cytokine secretion, and augmented memory cell proliferation. Taken together, our study provides critical insight regarding the cellular mechanisms underlying the effect of SARS-CoV-2 infection on host immune cell metabolism, and highlights immunometabolism as a promising therapeutic target for COVID-19 treatment.

Abstract 4255: Concurrent KRAS and TP53 mutations in pancreatic cancer real-world data (RWD) highlight convergent tumor evolutionary patterns

Abstract Pancreatic cancer is one of the most fatal of all cancers. Pancreatic adenocarcinomas have been historically difficult to treat, in part due to lack of early detection, and due to their resistance to conventional chemotherapy. There has recently been an improved understanding of the biologic processes driving pancreatic tumorigenesis, and an increase in clinical trial options available to patients, especially in the context of KRAS mutation positive pancreatic ductal adenocarcinoma (PDAC). KRAS is a known driver of pancreatic cancer initiation, with concurrent loss of TP53 in many cases. Among all mutated genes in pancreatic cancer, KRAS, TP53, CDKN2A, and SMAD4 are most frequently reported, followed by CDKN2B and ARID1A among others. Mutations in TP53 commonly found in pancreatic cancer are either missense or truncating. The specific type of mutation may be important to note since point mutations may produce dysfunctional proteins that acquire gain of functions that are tumor-promoting, although the most well-agreed upon effects of TP53 mutations stem from the absence or decrease of its normal physiologic functions including cell cycle arrest, apoptosis promotion, DNA damage response triggering, maintenance of cell polarity, and maintenance of genomic stability. Here we present an in-depth multi-dimensional analysis of 29 pancreatic adenocarcinoma patients, consisting of 11 patients with KRAS G12D mutations, 5 patients with KRAS G12V mutations, 3 patients with non-canonical KRAS mutations, and 10 patients with no KRAS mutations identified in their tumors. Individual TP53 mutations were highlighted in the context of associated KRAS mutations and tumor genomic landscape at the time of testing. Real-world data was collected with patient consent through community embedded Oncology Clinical Team (ceOCT) facilitated engagement. Tumor evolution prediction rules were constructed based on mutational landscapes of the 29 patients and evidence published in peer-reviewed journals. Pancreatic cancer patient data from the AACR GENIE Cohort v12.0-public database and the cBioportal for Cancer Genomics public site was subsequently used for testing predicted tumor evolutionary trajectories. Here we describe multiple independent combinations of KRAS and TP53 mutations leading to convergent functional outcomes for tumor cells, in the context of patient-specific gene mutation landscapes. Recurrent patterns of paired mutations with common cellular function dysregulation outcomes highlight core underlying mechanisms of pancreatic tumor evolution. Citation Format: Kaumudi Bhawe, Adrienne Nugent, Deb Christensen, Lauren Levine, Emma Shtivelman. Concurrent KRAS and TP53 mutations in pancreatic cancer real-world data (RWD) highlight convergent tumor evolutionary patterns. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4255.